Acute myeloid leukemia (AML) is the most common acute leukemia in adults, with 19,000 new cases annually in the US. A deoxycytidine nucleoside analog Cytarabine (AraC) is the principal chemotherapeutic agent used in AML. AraC-containing chemotherapeutic regimens induce complete remission in 65?80% of newly diagnosed AML patients, but a substantial minority of leukemias is intrinsically resistant to AraC. Furthermore, most of the initially responding patients relapse with resistant disease and poor response to subsequent therapies. As a result, the current overall survival rate for adult AML patients is less than 30%. The known mechanisms of AraC resistance involve changes in gene expression, at least some of which can be induced epigenetically by treatment with AraC or other chemotherapeutic drugs. A major cause of clinical resistance in AML is the presence of the leukemic stem cell population, resistant to conventional therapies. Novel therapies with the abilities to prevent the emergence of drug resistance or to target the leukemic stem cells are needed to improve patient outcomes. This application proposes a potentially transformative approach to leukemia therapy based on combining AraC with a novel class of preclinical agents that target transcription-regulating kinase CDK8 and its paralog CDK19. CDK8/19 inhibitors are characterized by a lack of toxicity, chemopotentiating activity, the ability to prevent transcriptional changes leading to drug resistance, inhibition of stem cell differentiation and proliferation and direct suppression of a subset of leukemias. We have found that the addition of CDK8/19 inhibitors to AraC drastically improves its long-term efficacy in human AML cell lines that do not respond to CDK8/19 inhibitors alone and inhibits the development of AraC resistance. A combination of CDK8/19 inhibitors with AraC also shows efficacy against AML lines with either acquired or intrinsic AraC resistance. Senex Biotechnology has developed a highly selective inhibitor of CDK8/19, Senexin B, and we now propose a program aimed at developing Senexin B/AraC combinations for AML therapy. In Phase I of this program, we will investigate the long-term in vitro efficacy of Senexin B/AraC combinations against a panel of human AML cell lines and determine if the synergistic effect is associated with the impact of CDK8/19 inhibition on the stem cell-like subpopulation of leukemic cells. To establish the conditions for efficient and sustained suppression of AML growth in vivo by Senexin B/AraC combination, we will first carry out pharmacokinetics and toxicology studies of Senexin B combined with AraC in immunodeficient mice. We will then test the efficacy of Senexin B alone in a mouse xenograft model of a Senexin B-sensitive human leukemia and of Senexin B/AraC combination in both Senexin B-sensitive and Senexin B-resistant mouse xenograft AML models. In the future Phase II, we will extend our studies to patient-derived leukemia samples and generate an IND package to initiate clinical studies of a CDK8/19 inhibitor/AraC combination for AML therapy.